Expression of inducible nitric oxide synthase activity in human colon epithelial cells: modulation by T lymphocyte derived cytokines (original) (raw)
Related papers
Free Radical Biology and Medicine, 2005
Inducible nitric oxide synthase (iNOS) activity in colonic epithelial HT-29 cells is modulated by the T-cell-derived cytokines IL-4 and IL-13, but is not affected by IL-10 despite its effect in models of colitis. We studied the effects of these cytokines on nitric oxide (NO) production by colonic tissue. IL-10 and IL-4 but not IL-13 suppressed the NO production and iNOS expression by inflamed tissue and cytokine-stimulated noninflamed tissue from patients with ulcerative colitis, whereas the three cytokines suppressed NO production in cytokine-stimulated biopsies from controls. To examine why colonic biopsies and HT-29 cells respond differently to immunomodulatory cytokines, a coculture of mixed mononuclear monocytes (MMC) and HT-29 cells was studied. Treatment of HT-29 cells with conditioned medium from IFN-g/LPS-stimulated MMC produced significant amounts of NO, which suggested the presence of an MMC-derived soluble factor modifying epithelial NO production. Pretreatment of IFN-g/LPS-stimulated MMC with IL-10 and IL-4 but not IL-13 suppressed NO production by HT-29 cells. Interestingly, pretreatment of HT-29 cells with IL-1 receptor antagonist suppressed the IFN-g/LPS-stimulated MMC-induced NO production. These results suggest that immunomodulatory cytokines might exert an inhibitory effect on NO up-regulation by colonic epithelium via the inhibition of MMC-derived soluble mediators, such as IL-1. D
Induction of iNOS in a rat model of acute colitis
1999
Induction of colitis by 2,4,6-Trinitrobenzenesulphonic acid (TNB) in the rat is a widely used experimental model of inflammatory bowel disease. Action of TNB as a hapten, induces colitis involving infiltration of colonic mucosa by neutrophils and macrophages and increased production of inflammatory mediators. The aim of the present study was to measure nitric oxide synthase (NOS) activity and characterize relations between inducible NOS (iNOS) activity and other signs of inflammation in TNB-induced colitis. A profound and sustained increase in the activity of iNOS was found in the colon. The activity of NOS in the spleen was also increased, but remained at low levels as compared to those in colon. No increases in plasma nitrite + nitrate concentrations were found suggesting local rather than systemic induction of iNOS. The increase in iNOS activity in the colon was preceded by macroscopic inflammatory lesions, like hyperemia, ulcerations and edema formation as well as neutrophil accumulation in the gastric mucosa and increased circulating concentrations of PGE2 metabolite (PGEM). Concentrations of PGEM in the plasma and myeloperoxidase activity (MPO; marker of neutrophil infiltration) in the gut declined in 48h whereas increased iNOS activity and the macroscopic inflammatory lesions remained over the 72h follow-up period. The results demonstrate increased local iNOS activity in TNB-induced colitis mimicking the situation in human inflammatory bowel disease.
Role of Blood- and Tissue-Associated Inducible Nitric-Oxide Synthase in Colonic Inflammation
The American Journal of Pathology, 2007
There is evidence that inducible nitric-oxide synthase (iNOS)-derived NO contributes to the pathophysiology of intestinal inflammation. The aims of this study were to assess the role of iNOS in the development of dextran sodium sulfate (DSS)-induced colonic inflammation and to define the contribution of tissue-specific iNOS expression to this inflammatory response. Study groups included: 1) wild-type (WT) mice; 2) WTfWT bone marrow chimeras with normal iNOS function; 3) WTfiNOS ؊/؊ chimeras (with functional blood cell iNOS, but iNOS-deficient tissue); 4) iNOS ؊/؊ fWT chimeras (with iNOS-deficient blood cells, but normal tissue iNOS activity); and 5) iNOSdeficient mice. In WT mice and WTfWT chimeras, DSS-induced colonic inflammation was characterized by bloody diarrhea and a high disease activity index. However, WTfiNOS ؊/؊ and iNOS ؊/؊ fWT chimeras and iNOS ؊/؊ mice exhibited an attenuated disease activity index, with parallel changes in histopathology. Colonic myeloperoxidase (MPO) was comparably elevated in DSS-treated WT mice (30.1 ؎ 1.7) and WTfWT chimeras (29.0 ؎ 1), whereas MPO was significantly reduced in iNOS ؊/؊ mice and iNOS ؊/؊ fWT chimeras (9.5 ؎ 1.7 and 15.6 ؎ 2.2, respectively). WTfiNOS ؊/؊ chimeras exhibited the lowest MPO activity (3.7 ؎ 0.6). Our findings implicate both blood cell-and tissue-derived iNOS in DSS-induced colonic inflammation, with tissue-associated iNOS making a larger contribution to the recruitment of inflammatory cells.
Pathophysiological role of nitric oxide in rat experimental colitis
International Journal of Immunopharmacology, 1998
Overproduction of nitric oxide "NO# by inducible nitric oxide synthase "iNOS# may contribute to the pathophysiology of ulcerative colitis[ A 1\3\5!trinitrobenzenesulfonic acid sodium salt "TNBS# colitis model was established to examine the e}ect of selective iNOS inhibition\ by S!"1!aminoethyl# isothiouronium bromide "ITU#\ on colonic mucosal cell damage and in~ammation[ Rats\ killed 6 days after TNBS\ had increased colonic mucosal levels of iNOS and interleukin!7 "IL!7#\ in addition to severe colonic in~ammation which was characterized by signi_cantly increased colon weight\ damage score and colonic myeloperoxidase activity "MPO# "a marker of neutrophil in~ux#[ TNBS!treated rats had markedly decreased body weight and thymus weight[ Administration of colitic rats with ITU signi_cantly inhibited iNOS activity:expression and tended to reduce mucosal levels of IL!7\ but no e}ect on MPO activity was observed[ Following ITU therapy\ colitic rats had reduced colonic damage and losses in body weight and thymus weight were reversed[ Improvement of TNBS colitis by ITU suggested that excess NO\ produced by iNOS\ may have contributed to the initiation: ampli_cation of colonic disease\ by mechanisms including enhancement of IL!7 release NO!mediated enhancement of pro!in~ammatory cytokine release was further investigated in vitro[ Lipo! polysaccharide "LPS# and interferon!g "IFN!g# stimulated release of nitrite\ lactate dehydrogenase "LDH#\ TNFa\ IL!0b and IL!7 from rat peritoneal macrophages\ all of which were signi_cantly reduced by ITU[ This suggests that NO!mediated cell damage enhances pro!in~ammatory mediator release from macrophages[ In addition\ enhancement of IL!7 and TNFa release was also partially NO!dependent in activated peritoneal neutrophils[ Therefore\ the amelioration of TNBS colitis by ITU could include inhibition of NO!mediated pro!in~ammatory cytokine release[ Þ 0887 International Society for Immunopharmacology[ Keywords] colitis\ trinitrobenzene sulfonic acid\ nitric oxide\ inducible nitric oxide synthase\ tumour necrosis factor\ interleukin!0\ interleukin!7
The Journal of Pathology, 1998
Nitric oxide (NO) and reactive oxygen species (ROS) are important mediators in the pathogenesis of inflammatory bowel disease (IBD). NO in IBD can either be harmful or protective. NO can react with superoxide anions (O 2 • • • • •-) yielding the toxic oxidizing agent peroxynitrite (ONOO -). Peroxynitrite induces nitration of tyrosine residues (nitrotyrosine) leading to changes of protein structure and function. The aim of this study was to identify the cellular source of inducible nitric oxide synthase (iNOS), and to localise superoxide anion producing cells in mucosal biopsies from patients with active IBD. Additional studies were performed to look at nitrotyrosine formation as a measure of peroxynitrite mediated tissue damage. For this antibodies against iNOS, eNOS, and nitrotyrosine were used. ROS producing cells were detected cytochemically. Inflamed mucosa of patients with active IBD showed intense iNOS staining in the epithelial cells. iNOS could not be detected in non-inflamed mucosa of IBD patients and control subjects. eNOS was present in blood vessels, without any difference in staining intensity between IBD patients and control subjects. ROS producing cells were increased in the lamina propria of IBD patients, a fraction of these cells were CD 15 positive. Nitrotyrosine formation was found on ROS positive cells. These results show that iNOS is induced in epithelial cells from patients with active ulcerative colitis or Crohn's disease. Nitration of proteins was only detected in the ROS producing cells at some distance from the iNOS producing epithelial cells. These findings indicate that tissue damage during active inflammation in IBD patients is probably more related to ROS producing cells than to NO. One may speculate that NO has a protective role when during active inflammation other mucosal defence systems are impaired.
Scandinavian Journal of Gastroenterology, 2005
Objective. The role of nitric oxide (NO) in the pathophysiology of inflammatory bowel disease (IBD) is controversial. The aim of this study was to investigate the expression and localization of nitric oxide synthase isoforms (iNOS, eNOS) in IBD colonic mucosa. Material and methods. Forty-four patients with IBD (24 ulcerative colitis (UC), 20 Crohn's disease (CD) and 16 controls) were investigated by colonoscopy. iNOS and eNOS in tissue sections was demonstrated by histochemistry (NADPH-diaphorase reaction) and immunohistochemistry. Cell type analysis and quantitative assessment of the iNOS immunoreactive (IR) cells and densitometry of iNOS in immunoblots were also performed. Results. iNOS-IR cells were significantly numerous in inflamed mucosa of UC (649/4 cells/mm 2 ) than in CD (49/2 cells/mm 2 ). iNOS-IR/ CD15-IR cells showed significant elevation in inflamed (i) versus uninflamed (u) UC mucosa (UC u 89/3%, UC i 859/10%) In CD, the percentage of iNOS-IR/CD68-IR cells was lower in inflamed sites (CD u 239/8%, CD i 49/3%). Immunoblot of biopsies revealed significant elevation of iNOS in active UC compared with uninflamed sites, whereas in CD no significant changes were detected. Differences were observed in eNOS and endothelial marker CD31 immunoreactivity. In patients with UC and in controls the ratios of eNOS/CD31-IR vessels were 82.3% and 92.0% respectively, whereas in CD the ratio was 8.3% with a concomitantly significant increase of CD31-IR vessels. The distribution and morphological characteristics of the NOS-IR inflammatory cells and endothelia were similar to those showing NADPH-diaphorase reactivity. Conclusions. Differences observed in the expression and distribution of NOS isoforms in immune and endothelial cells may contribute to better understanding of the structural and physiological changes in UC and CD.
Inducible nitric oxide synthase as a potential blood-based biomarker in inflammatory bowel diseases
PubMed, 2018
Aim: Here, we evaluated the role of (iNOS) as a blood-based biomarker of inflammatory bowel diseases (IBD). Background: Up-regulation of inducible nitric oxide synthase (iNOS) in the intestinal epithelial cells has been closely associated with the initiation and maintenance of intestinal inflammation in IBD. Methods: In a case-control design, 59 IBD patients and 30 healthy control subjects were participated in this study. A total of 10 ml blood sample was taken from each participant. Blood leukocytes were isolated and iNOS mRNA expression level was evaluated in the isolated leukocytes using relative quantitative Real-time PCR. Results: The patients' population included 40 ulcerative colitis (UC) and 19 Crohn's disease (CD) patients. The flare and remission phase of disease were seen in 43 and 16 patients, respectively. The mean iNOS mRNA expression was not significantly different between the IBD patients and healthy controls (p=0.056). The mean iNOS mRNA expression was significantly higher in the flare phase of the disease compared to the remission phase (p=0.039). No significant difference was observed between the mean iNOS mRNA expression in the blood leukocytes of UC and CD patients (p=0.82). Conclusion: iNOS is differently expressed in the blood leukocytes of active vs. inactive IBD disease. Thus, it could be potentially used as a non-invasive blood-based biomarker of IBD.
Nitric oxide in inflammatory bowel disease: a universal messenger in an unsolved puzzle
Immunology, 2004
In recent years, nitric oxide (NO), a gas previously considered to be a potentially toxic chemical, has been established as a diffusible universal messenger that mediates cell-cell communication throughout the body. Constitutive and inducible NO production regulate numerous essential functions of the gastrointestinal mucosa, such as maintenance of adequate perfusion, regulation of microvascular and epithelial permeability, and regulation of the immune response. Up-regulation of the production of NO via expression of inducible nitric oxide synthase (iNOS) represents part of a prompt intestinal antibacterial response; however, NO has also been associated with the initiation and maintenance of inflammation in human inflammatory bowel disease (IBD). Recent studies on animal models of experimental IBD have shown that constitutive and inducible NO production seems to be beneficial during acute colitis, but sustained up-regulation of NO is detrimental. This fact is also supported by studies on mice genetically deficient in various NOS isoforms. However, the mechanism by which NO proceeds from being an indispensable homeostatic regulator to a harmful destructor remains unknown. Furthermore, extrapolation of data from animal colitis models to human IBD is questionable. The purpose of this review is to update our knowledge about the role of this universal mediator and the enzymes that generate it in the pathogenesis of IBD.